摘要
目的:观察含铈硅酸钙(Ce-CS)修饰的PET人工韧带对大鼠骨髓间充质干细胞(rat bone marrow mesenchymal stem cells,rBMSCs)增殖和成骨分化的影响,及其在动物体内对移植物-骨愈合的影响。方法:根据PET韧带表面处理情况分为2组:PET组(PET材料未经任何表面处理)和Ce-CS组(PET韧带经含铈硅酸钙进行表面修饰)。将rBMSCs接种到上述两组不同的材料上,培养一段时间后进行各项检测。扫描电镜(SEM)和X线能谱分析(EDS)观察材料涂层情况。rBMSCs在材料上接种培养1、3、7天后,使用Cell Counting Kit-8(CCK-8)试剂盒,检测rBMSCs细胞增殖情况。rBMSCs经成骨诱导培养7天和14天后,检测碱性磷酸酶(ALP)的活性,Real Time PCR(RT-PCR)检测骨钙蛋白(OCN)、Ⅰ型胶原(COL-1)的表达;诱导培养21天后,进行茜素红染色。选取12只新西兰大白兔,建立关节外腱-骨愈合模型。左侧植入未经处理的PET韧带,右侧植入Ce-CS修饰的PET韧带。在韧带植入8周后处死兔子,取出股骨样本,分别进行Micro-CT和Masson、HE染色。结果:SEM和EDS结果显示,Ce-CS材料涂层的PET韧带制备成功。CCK-8结果显示,Ce-CS组细胞在接种培养后第3天和第7天的增殖情况要优于对照组(P<0.05)。诱导培养7天和14天后,Ce-CS组ALP活性均高于对照组(P<0.05);诱导培养7天后OCN表达高于对照组(P<0.05);诱导培养14天后,OCN、COL-1表达均高于对照组(P<0.05)。Ce-CS组形成的矿化结节要多于对照组(P<0.05)。Micro-CT结果显示,CE-CS组骨缺损处的骨矿密度(BMD)、相对骨体积(BV/TV)均高于对照组。Masson、HE染色显示Ce-CS组韧带与宿主骨之间连接较为紧密,有较多新生骨组织生成,对照组与宿主骨之间连接疏松,未见有新生骨组织生成。结论:含铈硅酸钙修饰的PET韧带在体外能够明显促进大鼠骨髓间充质干细胞的增殖和成骨分化,体内能够促进新骨再生,促进人工韧带与宿主骨之间的愈合。
Objective To observe the effect of cerium incorporated-calcium silicate modified polyethylene terephthalate(PET)on the proliferation and osteogenic differentiation of rat bone marrow mesenchymal stem cells(rBMSCs)and the graft-to-bone healing in vivo.Methods The PET sheets were divided into two different groups according to the surface treatment,the PET group(PET sheets without any surface treatment)and the Ce-CS group(PET sheets modified with cerium incorporated-calcium silicate).The rBMSCs were seeded onto the above two different PET sheets,and a series of tests were carried out after culturing.The scanning electron microscopy(SEM)and X-ray spectroscopy(EDS)were used to detect the coating on the surface of PET sheets.The proliferation of rBMSCs on the surface of PET sheets were measured using the cell counting kit-8(CCK-8)assay after 1,3,and 7 days of incubation.After 7 days and 14 days of induction in osteogenic induction medium,the activity of alkaline phosphatase(ALP)were assayed.The real-time polymerase chain reaction(RT-PCR)were carried out after 7 days’and 14 days’treatment with osteogenic induction medium to evaluate the relative expression of osteogenic genes,such as osteocalcin(OCN)and collagen type I(COL1).The alizarin red staining was employed 21 days after culturing to assess the mineralization of rBMSCs.Twelve male New Zealand white rabbits were enrolled in the study and an extra-articular graft-bone healing model was established by implanting the PET sheets into the distal femur.The Ce-CS modified PET were implanted into the right femur while the untreated PET sheets were implanted into the right.Rabbits were sacrificed 8 weeks after implantation and femurs were harvested for Micro-CT,Masson,and HE staining.Results The SEM and EDS results showed that Ce-CS was successfully coated onto the surface of PET sheets.The proliferation of rBMSCs of the Ce-CS group was superior to the PET group on the 3 rd and 7 th days of incubation(P<0.05).The ALP activity of rBMSCs of the Ce-CS group on the 7 th and 14 th days of induction was significantly higher than that of controls(P<0.05).The relative expression level of OCN(P<0.05),as well as COL-1(P<0.05),was improved in the Ce-CS group on the 14 th days compared to the control group.The number of mineralized nodules in the Ce-CS group was more than that in the control group(P<0.05).The Micro-CT images showed that the bone mineral density(BMD)and relative bone volume(BV/TV)in the right bone defect was higher than that in the left side.Masson and HE staining images showed that new bone was formed between the Ce-CS coated PET ligaments and the host bone.There was a loose connection between the untreated ligaments and the host bone and much amount of fibrous tissue between them.Conclusions The Ce-CS coated PET can significantly enhance the proliferation and osteogenic differentiation,and promote the bone regeneration in vivo,bettering the graft-to-bone healing.
作者
康展荣
傅声扬
俞斌
丁惠锋
胡健
禹宝庆
黄建明
Kang Zhanrong;Fu Shengyang;Yu Bin;Ding Huifeng;Hu Jian;Yu Baoqing;Huang Jianming(Department of Orthopaedics,Shanghai Pudong Hospital Affiliated to Fudan University,Shanghai 201399,China;School of Material Science and Engineering,University of Shanghai for Science and Technology,Shanghai 200093,China;Department of Orthopaedics,Shanghai Public Health Clinical Center,Shanghai Medical School,Fudan University,Shanghai 200032,China)
出处
《中国运动医学杂志》
CAS
CSCD
北大核心
2019年第5期396-405,共10页
Chinese Journal of Sports Medicine
基金
上海市浦东新区卫生系统学科带头人培养计划资助(PWRd2017-03)
上海市浦东医院人才培养计划(PX2018-02)
上海市浦东新区医学学科建设项目(PWYgy2018-09)
关键词
聚对苯二甲酸乙二醇酯
铈
硅酸钙
骨髓间充质干细胞
人工韧带
移植物-骨愈合
polyethylene terephthalate
cerium
calcium silicate
bone marrow mesenchymal stem cells
artificial ligament
graft-to-bone healing
作者简介
同等贡献作者:康展荣,Email:zrkang2016@163.com;同等贡献作者:傅声扬,Email:fushengyang@sina.com;通信作者:黄建明,Email:hjm19611009@163.com
